This application is for an R24 Consortium Grant (PA-00-099) to support collaborative research between five independent groups at four academic institutions. The long-term goals of this research are to more fully understand the cooperative functioning of the protein complexes that carry out respiration, and to infer how these functions may have changed during human evolution. The major hypothesis being tested is that many of the respiratory chain proteins, which are encoded both by the nuclear and mitochondrial genomes, have adaptively co-evolved during primate evolution. Previous studies have suggested that certain subunits of the electron transport system (ETS) underwent episodes of positive Darwinian selection and co-evolution on ancestral lineages leading to the anthropoid primates (monkeys, apes, and humans). The consortium will refine these studies, both by sequencing ETS genes from a dense and diverse phylogenetic sample of primates and by applying rigorous maximum likelihood (ML) methods of analysis. These approaches will be applied to all of the ETS complexes, with most emphasis on those for which crystal structures have been determined. These goals will be accomplished through the following Specific Aims: Specific Aim 1: Mitochondrial DNA (mtDNA) genomes will be sequenced from phylogenetically-diverse primates. The protein-coding genes, which code for some of the ETS subunits, will be individually analyzed for evidence of positive Darwinian selection. The whole genomes will be analyzed phylogenetically. Specific Aim 2: Selected nuclear genes that code for proteins that interact with the mtDNA-encoded subunits will be sequenced from the same species. These genes will be analyzed for evidence of positive selection. Specific Aim 3: The interacting proteins, and the genes that encode them, will be analyzed for evidence for adaptive co-evolution by phylogeny-based ML methods that incorporate tertiary structural information. These data and analyses will be used to more fully test the hypothesis that certain ETS proteins underwent episodes of positive Darwinian selection on ancestral primate lineages leading to humans. This information has important implications for understanding energy metabolism in humans, particularly as related to brain expansion during human evolution. Furthermore, information about which proteins, or parts of proteins, have undergone recent positive selection may help define targets for intervention in certain human mitochondrial diseases.